Dating the origin of the CCR5-Delta32 AIDS-resistance allele by the coalescence of haplotypes.

The CCR5-Delta32 deletion obliterates the CCR5 chemokine and the human immunodeficiency virus (HIV)-1 coreceptor on lymphoid cells, leading to strong resistance against HIV-1 infection and AIDS. A genotype survey of 4,166 individuals revealed a cline of CCR5-Delta32 allele frequencies of 0%-14% across Eurasia, whereas the variant is absent among native African, American Indian, and East Asian ethnic groups. Haplotype analysis of 192 Caucasian chromosomes revealed strong linkage disequilibrium between CCR5 and two microsatellite loci. By use of coalescence theory to interpret modern haplotype genealogy, we estimate the origin of the CCR5-Delta32-containing ancestral haplotype to be approximately 700 years ago, with an estimated range of 275-1,875 years. The geographic cline of CCR5-Delta32 frequencies and its recent emergence are consistent with a historic strong selective event (e.g. , an epidemic of a pathogen that, like HIV-1, utilizes CCR5), driving its frequency upward in ancestral Caucasian populations.     

Inter-simple-sequence-repeat (ISSR) polymorphisms are useful for finding markers associated with disease resistance gene clusters

 We describe a simple and new approach, based on inter-simple sequence repeats (ISSRs), for finding markers linked to clusters of disease resistance genes. In this approach, simple sequence repeats (SSR) are used directly in PCR reactions, and markers found to be linked to disease resistance genes provide important information for the selection of other sequences which can be used with PCR to find other linked markers. Based on an ISSR marker linked to a gene of interest, many new markers can be identified in the same region. We previously demonstrated that ISSR markers are useful in gene tagging and identified a marker, UBC-855₅₀₀, linked to the gene for resistance to fusarium wilt race 4 in chickpea. This ISSR marker provided the information used in the present study for selecting other primers which amplified a region linked to the gene for resistance to fusarium wilt race 4. The primers were based on homology with the (AC)_n sequence and were used for PCR amplifications. Changes in the sequence were at the anchor region of the primers. The repeat (AC)₈T amplified a marker, UBC-825₁₂₀₀, which was located 5.0 cM from the gene for resistance to fusarium wilt race 4 and was closer than other markers. These results indicated that ISSR markers can provide important information for the design of other primers and that by making changes at the 3′ and 5′ anchors close linkage to the desired gene can be found. The approach allows rapid scanning of the targeted region and may provide important information for genome analysis of plant species. Received: 20 January 1998 / Accepted: 19 March 1998     

Simple sequence repeats for the genetic analysis of apple

 The development of highly informative markers, such as simple sequence repeats, for tagging genes controlling agronomic characters is essential for apple breeding. Furthermore the use of these markers is fundamental both for variety identification and for the characterisation and management of genetic resources. We have developed 16 reliable simple sequence repeat (SSR) markers that amplify all alleles from a panel of 19 Malus x domestica (Borkh.) cultivars or breeding selections and from Malus floribunda 821. Those markers show a high level of genetic polymorphism, with on average 8.2 alleles per locus and an average heterozygosity of 0.78. Due to this high level of polymorphism, it was possible using two selected SSRs to distinguish all cultivars except Starking and Red Delicious. Ten of the markers we developed have been mapped on a RAPD linkage map, proving their Mendelian segregation as well as their random distribution in the apple genome. Finally, we discuss the importance of using co-dominant markers in outbreeding species. Received: 8 October 1997 / Accepted: 9 December 1997     

The use of microsatellite markers for detection of genetic diversity in barley populations

 A barley lambda-phage library was screened with (GA)n and (GT)n probes for developing microsatellite markers. The number of repeats ranged from 2 to 58 for GA and from 2 to 24 for GT. Fifteen selected microsatellite markers were highly polymorphic for barley. These microsatellite markers were used to estimate the genetic diversity among 163 barley genotypes chosen from the collection of the IPK Genebank, Germany. A total of 130 alleles were detected by 15 barley microsatellite markers. The number of alleles per microsatellite marker varied from 5 to 15. On average 8.6 alleles per locus were observed. Except for GMS004 all other barley microsatellite markers showed on average a high value of gene diversity ranging from 0.64 to 0.88. The mean value of gene diversity in the wild forms and landraces was 0.74, and even among the cultivars the gene diversity ranged from 0.30 to 0.86 with a mean of 0.72. No significant differences in polymorphism were detected by the GA and GT microsatellite markers. The estimated genetic distances revealed by the microsatellite markers were, on average , 0.75 for the wild forms, 0.72 for landraces and 0.70 among cultivars. The microsatellite markers were able to distinguish between different barley genotypes. The high degree of polymorphisms of microsatellite markers allows a rapid and efficient identification of barley genotypes. Received: 26 November 1997 / Accepted: 19 January 1998     

Development,characterization and utilization of genomic microsatellite markers in turmeric (Curcuma longa L.)

Development of a robust set of 18 genomic microsatellite markers from turmeric (Curcuma longa L.) and its effective utilization in estimating the genetic diversity of 20 turmeric accessions are described. A total of 103 alleles were detected with an average of 5.7 alleles per locus. These markers displayed varied levels of polymorphism as evident from its discriminating power ranging from 0.19 to 0.70. The UPGMA cluster analysis of genetic distance values resolved the 20 turmeric accessions into five main groups. Three sets of genetically identical accessions were detected within the analyzed accessions, suggesting a revisit of the germplasm collection strategy based on vernacular identity. The entire grouping pattern of the entities was loose and independent of their geographical origins. These polymorphic SSR markers would be useful for the population genetic studies and germplasm management of turmeric.     

Assessment of the genetic diversity among Glyptosternum maculatum, an endemic fish of Yarlung Zangbo River, Tibet, China using SSR markers

The genetic diversity of Glyptosternum maculatum populations from Nyang River, Lhasa River, and Shetongmon Reach of Yarlung Zangbo River was assessed using six microsatellite markers. Overall, the genetic diversity across the three populations was low. The Shetongmon population exhibited the highest level of genetic diversity in terms of number of alleles and effective alleles, heterozygosity, and polymorphic information content value, followed by the Nyang population and Lhasa population. The analysis of molecular variance demonstrated that almost the variation (86.64%) occurred within populations. The differentiation among populations was not significant, and population structure was weak. These results revealed that three natural populations of G. maculatum are not genetically differentiated and the large disparity of living altitude did not caused genetic differentiation between different populations. Our observations will help identify the genetic relationship among populations to understand the genetic diversity of G. maculatum.     

The distribution of coalescence times and distances between microsatellite alleles with changing effective population size

We investigate the effects of past changes of the effective population size on the present allelic diversity at a microsatellite marker locus. We first derive the analytical expression of the generating function of the joint probabilities of the time to the Most Recent Common Ancestor for a pair of alleles and of their distance (the difference in allele size). We give analytical solutions in the case of constant population size and the geometrical mutation model. Otherwise, numerical inversion allows the distributions to be calculated in general cases. The effects of population expansion or decrease and the possibility to detect an ancient bottleneck are discussed. The method is extended to samples of three and four alleles, which allows investigating the covariance structure of the frequencies f(k) of pairs of alleles with a size difference of k motifs, and suggesting some approaches to the estimation of past demography.     

Development of a microsatellite marker set applicable to genome-wide screening of cynomolgus monkeys (<Emphasis Type="Italic">Macaca fascicularis</Emphasis>)

To develop a microsatellite marker set applicable to genome-wide screening of cynomolgus monkeys (Macaca fascicularis), 148 microsatellite markers were selected from the human genome database. The polymorphisms and inheritance of PCR products were determined by screening twenty unrelated monkeys and by analysis of three families, respectively. As a result, 106 primers (72%) gave PCR products of the size expected for humans and rhesus monkeys. Among these products, polymorphism and single-gene inheritance in cynomolgus monkeys was observed for 66 markers (62%). The average number of alleles at the 66 polymorphic loci was 5.86 (range 2–10), and average heterozygosity was 0.63 (range 0.10–0.88). This is the first report of microsatellite markers for cynomolgus monkeys. Chromosomal mapping of these markers is now in progress.     

Mapping of Fhb2 on chromosome 6BS: a gene controlling Fusarium head blight field resistance in bread wheat (Triticum aestivum L.)

Fusarium head blight (FHB) is one of the most important fungal wheat diseases worldwide. Understanding the genetics of FHB resistance is key to facilitate the introgression of different FHB resistance genes into adapted wheat. The objective of this project was to study the FHB resistance QTL on chromosome 6B, quantify the phenotypic variation, and qualitatively map the resistance gene as a Mendelian factor. The FHB resistant parent BW278 (AC Domain*2/Sumai 3) was used as the source of the resistance allele. A large recombinant inbred line (RIL) mapping population was developed from the cross BW278/AC Foremost. The population segregated for three known FHB resistance QTL located on chromosomes 3BSc, 5A, and 6B. Molecular markers on chromosome 6B (WMC104, WMC397, GWM219), 5A (GWM154, GWM304, WMC415), and 3BS (WMC78, GWM566, WMC527) were amplified on approximately 1,440 F_2:7 RILs. The marker information was used to select 89 RILs that were fixed homozygous susceptible for the 3BSc and 5A FHB QTLs and were recombinant in the 6B interval. Disease response was evaluated on 89 RILs and parental checks in the greenhouse and field nurseries. Dual floret injection (DFI) was used in greenhouse trials to evaluate disease severity (DS). Macroconidial spray inoculations were used in field nurseries conducted at two locations in southern Manitoba (Carman and Glenlea) over two years 2003 and 2004, to evaluate disease incidence, disease severity, visual rating index, and Fusarium-damaged kernels. The phenotypic distribution for all five-disease infection measurements was bimodal, with lines resembling either the resistant or susceptible checks and parents. All of the four field traits for FHB resistance mapped qualitatively to a coincident position on chromosome 6BS, flanked by GWM133 and GWM644, and is named Fhb2. The greenhouse-DS trait mapped 2 cM distal to Fhb2. Qualitative mapping of Fhb2 in wheat provides tightly linked markers that can reduce linkage drag associated with marker assisted selection of Fhb2 and aid the pyramiding of different resistance loci for wheat improvement.     

The genetic structure of fermentative vineyard-associated <Emphasis Type="Italic">Saccharomyces cerevisiae</Emphasis> populations revealed by microsatellite analysis

From the analysis of six polymorphic microsatellite loci performed in 361 Saccharomyces cerevisiae isolates, 93 alleles were identified, 52 of them being described for the first time. All these isolates have a distinct mtDNA RFLP pattern. They are derived from a pool of 1620 isolates obtained from spontaneous fermentations of grapes collected in three vineyards of the Vinho Verde Region in Portugal, during the 2001–2003 harvest seasons. For all loci analyzed, observed heterozygosity was 3–4 times lower than the expected value supposing a Hardy–Weinberg equilibrium (random mating and no evolutionary mechanisms acting), indicating a clonal structure and strong populational substructuring. Genetic differences among S. cerevisiae populations were apparent mainly from gradations in allele frequencies rather than from distinctive “diagnostic” genotypes, and the accumulation of small allele-frequency differences across six loci allowed the identification of population structures. Genetic differentiation in the same vineyard in consecutive years was of the same order of magnitude as the differences verified among the different vineyards. Correlation of genetic differentiation with the distance between sampling points within a vineyard suggested a pattern of isolation-by-distance, where genetic divergence in a vineyard increased with size. The continuous use of commercial yeasts has a limited influence on the autochthonous fermentative yeast population collected from grapes and may just slightly change populational structures of strains isolated from sites very close to the winery where they have been used. The present work is the first large-scale approach using microsatellite typing allowing a very fine resolution of indigenous S. cerevisiae populations isolated from vineyards.